Sperm strainer system

ABSTRACT

The invention in the simplest form is a passive method and apparatus for filtering motile sperm from a sperm sample. The invention passively filters the motile sperm using a nucleopore membrane that is contained within a membrane assembly, wherein the membrane assembly is disposed in a container. In operation, a medium is placed in the container so the membrane contacts the medium. A sperm sample is placed one side of the membrane assembly, and the motile sperm migrate through the membrane leaving the non-motile sperm behind, where they can be easily extracted.

CROSS REFERENCE TO RELATED APPLICATIONS

This application claims priority under 35 U.S.C. §119(e) from aprovisional patent Ser. No. 60/068,240 application filed on Dec. 19,1997, which is incorporated herein by reference for all purposes.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention is a method and apparatus that provides aninexpensive, quick, and portable means to separate motile sperm from asperm sample. The invention passively filters the motile sperm using anucleopore membrane.

2. Background Art

Artificial insemination has become a much more frequent procedure due toa variety of sociological, economic, and perhaps environmental reasons.The number of inseminations in the United States and worldwide has beenincreasing, and will likely continue to increase in the next millennium.A number of causes have been cited for this increase. The age ofchildbearing has increased, as people are delaying children andmarriage. The increased age lowers the probability of fertilization forboth men and women. More and more women are also raising children bythemselves without a partner, and electing artificial insemination as ameans of fertilization. In addition, those that would otherwise not beable to have children now have a medical option that is relativelyaffordable. An additional factor may be that the sperm count among maleshas been declining, making fertilization more difficult. And finally,environmental factors have been blamed for decreased fertility of bothmen and woman.

There are currently a wide array of artificial insemination methods suchas, intracervical, intrauterine (IUI), intratubular and directintraperitoneal (DIPI) insemination, gamete intrafallopian transfer(GIFT), in vitro fertilization and embryo transfer (IVFET), zygoteintrafallopian transfer (such as ZIFT, PROST and TET), peritoneal oocyteand sperm transfer (POST), and sex selection, among others. Astechnology advances, other methods are certain to follow, however,regardless of the process, high motile sperm are preferred.

As an example, the Intrauterine Insemination (IUI) and In Vitrofertilization (IVF) methods attempt to recreate the reproductive processby placing sperm and eggs together in an environment conducive tofertilization either in the womb or outside the womb. The fertilizationprocess requires the sperm to actively invade the egg and commencefertilization. Motile sperm are much more likely to penetrate the egg.

A typical semen sample contains materials such as paternal plasma,protein, leukocytes, spermdecapitation factors and other extraneousmaterials, and dead, agglutinated or nonviable spermatozoa. Thesematerials are known to interfere with successful fertilization, and withthe successful maintenance of a fertilized ovum in the female patient.As an example, seminal plasma can cause severe uterine cramping, and ina worst case result in spontaneous abortion of the fertilized ovum.Thus, it is desirable to remove those spermatozoa having relatively lowmotility or possessing unhealthy, damaged or abnormal membranes.

The total number of sperm in an ejaculate is a measure of fertility,however, the percentage of motile sperm is much more important,especially when considering alternate reproductive means.

Sperm are categorized according to the their exhibited motility, asexhibited by Table 1, below.

                  TABLE 1                                                         ______________________________________                                        Motility Index                                                                Degree of Motility                                                                       Type of Motility                                                   ______________________________________                                        0          No motility, or movement of tail with no forward                              progression                                                        1          20% or less showing forward progression (sluggish                             movement)                                                          2          20%-50% showing forward progression                                3          50%-80% showing forward progression                                4          80%-100% showing forward progression (very                                    rapid movement)                                                    ______________________________________                                    

The percentage or motile sperm showing progressive swimming movements isa measure of the fertility of the sperm sample. The higher thepercentage, the higher quality of the sperm sample, and the greater thelikelihood that the sample will achieve fertilization.

A high quality sperm sample is important for many reasons. The processof artificial insemination is not only costly economically, but ispsychologically expensive. Unsuccessful attempts have devastatingeffects on the patients.

Higher quality sperm samples are also important considerations when thesample is subject to freezing or aqueous dilution, because theseprocesses tend to kill or weaken the sample. Thus, only the highestquality sperm may survive the processing procedures to which the spermare subjected.

In summary, higher viability spermatozoa are more likely to lead tosuccessful fertilization and impregnation. The higher quality sperm arealso more likely to survive freezing (cryopreservation) and otherprocessing procedures.

In order to reduce the aforementioned problems, attempts have been madeto provide a simple, portable, and inexpensive method and device for thefiltration of sperm. However, the prior art devices and methods hadsignificant disadvantages and shortcomings.

Various methods of selecting the more active sperm have been utilized inthe past, such as the swim up. Swim down and Percoil density gradientcentrifugation techniques. Swim-up methods are commonly used to processfresh or frozen specimens for the IUI and IVF procedures. The sperm isplaced in a medium and subjected to a centrifuge process. The moremotile sperm swim to a level where they can be extracted. Such methodsemploy multiple tube and centrifugation steps that are time consumingand can lead to a low recovery of motile sperm.

Table 2 below, provides a comparison of some features of these methods.

                  TABLE 2                                                         ______________________________________                                        Characteristics of Prior Art Methods                                          Feature Swim-Up   Strain Ex Percoll Enhance-S Plus                            ______________________________________                                        Sperm   None      None      1       1 wash/                                   Preparation                 wash/centri-                                                                          centrifugation                            after                       fugation                                          liquefaction                                                                  Preparation                                                                           Prepare   None      Build a 2                                                                             Build a 2 step                            of device                                                                             buffer solu-        step gradient                                                                         gradient                                          tion, pour                                                                    into plate                                                            Sperm                                                                         Quality                                                                       Motility                                                                              Higher %  Higher %  Same    Same motility                                     motile    motile    motility                                          Viability                                                                             unknown   unknown   unknown unknown                                   Leukocyte                                                                             unknown   unknown   unknown unknown                                   contaminants                                                                  Washes re-                                                                            None      None      1-2     1-2                                       quired after                                                                  separation                                                                    Total # 1         1         2-3     2-3                                       Centrifu-                                                                     gation                                                                        Endotoxin                                                                             No        No        Yes     Yes                                       Contamina-                                                                    tion                                                                          ______________________________________                                    

One prior art device used for separating the motile sperm from a spermsample is the migration-sedimentation chamber. This system uses aculture medium with the sperm sample suspended in the medium, andrequires a centrifuge to rapidly spin the chamber to separate the motilesperm. And, because it is an active process, it increases the likelihoodthat motile sperm will be excluded from the sample set, or damagedduring the process, thus producing a lower recovery sample.

In operation, the sperm sample is placed in a migration-sedimentationchamber, preferably in the lower well portion. The chamber is subjectedto centrifugation, and the re-suspended sperm pellet is transferred tothe gallery segment of the chamber. The portion of sperm in the well isthe motile portion, and is used for insemination.

U.S. Pat. No. 5,185,246 ('246) is a method for semen analysis employinga membrane separation. This patent discloses a method to separate theparticulate (cells, spermatozoa, and other particulate) in the wholesemen from the seminal plasma which contains many soluble compounds,proteins, hormones, small molecules and electrolytes. The pore size isspecifically designed for such a separation, and all sperm cells areblocked from passing through the membrane. The driving force of the '246patent is a vacuum, and the sperm cells are stuck to the filter, deadand immobile, amenable only for in-situ testing or further disruption. Aspecific volume, preferably 100 microliters with various aqueousdilutions is employed. The separation apparatus of the '246 patent is awell filter plate that is used for multiple testing in the same round ofprocessing.

The '246 patent is related to tests of andrological significance thatneed be performed on either the seminal plasma (in the absence of spermcells) or on the surface of the sperm cells (in the absence of theseminal plasma). But, the cells in these cases need not be viable, neednot be motile, and need not be freely dispersed in an aqueous bufferedmedium for retrieval. The '246 patent discloses a device for providingeasier diagnostic access to specific components of the semen sample.

Another such invention is described in U.S. Pat. No. 5,575,914, whichdiscloses a conduit that has glass wool as a strainer mechanism. Theglass wool is compressed to a density sufficient to permit passage ofthe more motile sperm while blocking the lesser or non-motile sperm. Inthe preferred embodiment, the conduit is a nested pair of tubes that tryto keep the glass wool density consistent.

The invention of U.S. Pat. No. 4,009,260 describes the fractionation ofsperm through layered migration of different layers having differingdensities. The more motile sperm penetrate deeper into the layers.Similarly, U.S. Pat. No. 4,007,087 also discloses the fractionation vialayered solutions contacting the sperm sample.

U.S. Pat. No. 5,427,946 discloses a channeling apparatus, where thereare inlet ports, flow channels, and nesting chambers. The sperm sampleis applied at the inlet port, and only the motile sperm are capable ofreaching the chambers.

In summary, problems heretofore exhibited in the art include having topurchase and maintain additional equipment such as a centrifuge. Priormethods required taking trips to other sites to perform the separationor centrifuge process. Other methods required extensive time to separatethe motile sperm. In addition, prior art methods that used activeseparation schemes removed motile sperm and reduced the recovery lot.

What is needed is a method and apparatus that can passively isolatehigher viability spermatozoa from the spermatozoa having relativelylower motility. The device should be inexpensive, portable, andeasy-to-use. The method should be sufficient to screen out the lessersperm and other compounds, leaving only high quality sperm forinsemination. What is needed is an efficient device and method ofremoving the less motile sperm without costly or time-consumingprocedures, and this new system should provide a quick and inexpensiveway to increase the likelihood of fertilization.

SUMMARY OF THE INVENTION

The present invention has been made in consideration of theaforementioned background. It is therefore an object of the presentinvention to provide a system for conveniently, quickly, andinexpensively separating motile sperm from a sperm sample. Thisinvention most generally relates to a method and an apparatus ofstraining sperm. In particular, it relates to a method of straining andseparating motile sperm from a larger sample set without usingadditional machinery or devices.

One object of the present invention is to separate motile sperm from thenon-motile sperm or leukocytes.

Another object o f the invention is to provide an inexpensive and easyto use device, such that each patient can use a clean sperm filter.

The membrane of the present invention is large enough to allow for thepassage of sperm through the membrane. In the preferred embodiment, a5-8 micron pore size is utilized.

The driving impetus of the present invention is the ability of qualitysperm to move and swim through the membrane. The non-motile sperm andleukocytes move slightly, if at all, and therefore do not penetrate themembrane. No other force, such as a vacuum, suction, or gravity isrequired. Any artificially induced stress or pressure would defeat thepurpose of the present invention by forcing non-motile sperm through themembrane and also possibly damaging the motile sperm by the appliedforce. The natural, self-actuated migration of the more motile sperm inthe semen bath assures that the sperm that do encounter and work theirway through the membrane will have a significantly higher averagemotility index than the remaining sperm.

The present invention further provides a method and apparatus for theseparation of motile swimming cells from the dead/non-motile spermatozoaand from white blood cells that contaminate semen in case ofinflammatory events. The present invention ascribes no significance tothe separation of seminal plasma from the particulate matter since it isnot a desired component that needs special care and attention.

The method and apparatus of the present invention is for the separationof the `better` sperm, unattached to membranes or anything else, and ina form immediately available for retrieval and subsequent use inartificial insemination by means of a filtering membrane.

The motile sperm that penetrate the membrane may be supported in abuffered medium. After an incubation period, the motile sperm isimmediately ready for functional analysis and the various methods ofartificial insemination.

The cell strainer device in conjunction with the present methodologyoffers a rapid and efficient method for improving the sperm quality as aroutine sperm preparation for artificial insemination. This spermstrainer device is a one-step system that enables the recovery ofhigh-quality motile sperm in minimal time. No washing or centrifugationrequired, and the sperm preparation can be done in the office or at thedesired location.

The preferred embodiment of the present invention includes a cellstrainer unit with a 5 micron nylon mesh size and with an openable lid.This strainer is compatible with a 1 ml dish and has a screw cap. The 1ml dish is used to hold the medium and retain the cell strainer unitwhen properly engaged. A disposable plastic pipette is used to place thesperm sample into the cell strainer, and a 1 ml disposable syringe withneedle is used to place the medium into the dish and to extract thesperm sample when incubation is completed.

The method of operation of the preferred embodiment comprises placing 1ml of a medium in the container or dish using the syringe with theneedle. The user then puts 1-1.5 ml of liquefied semen into the straineror membrane unit using a disposable plastic pipette. The lid on thestrainer is closed, capturing the membrane or mesh securely in place,and the strainer unit is secured onto the dish. The unit is left for30-45 minutes for incubation at 37° C. in 5% CO₂. At the end of theincubation period, the user can remove the strainer unit from the dishand discard the cell strainer. The sperm sample remaining in the dish isthe motile sperm sample and the user can carefully aspirate all mediafrom the dish, using the syringe with needle. The recovered sperm inmedia is ready to use for artificial insemination. For IUI applications,it is necessary to remove the needle from the syringe and attach thecatheter and perform the IUI as normally done in the industry.

Tables 2, 3 and 4 below illustrate three actual examples of the originalsemen sample and the results of the Swim-Up Method of the prior art, ascompared to the Cell-Strainer Method.

                  TABLE 2                                                         ______________________________________                                        Original versus Separation Results for Patient XY                                                        Cell-Strainer                                                                          Cell-Strainer                                       Original                                                                              Swim-up  Method   Method                                    Patient XY                                                                              Semen   Method   (Vertical)                                                                             (Horizontal)                              ______________________________________                                        Volume (ml)                                                                             2.2     0.5      0.5      0.5                                       Sperm count/ml                                                                          52      35       34       29                                        (million)                                                                     Motility %                                                                              55      72       70       73                                        Progression                                                                             3       4        4        4                                         Velocity  29      33       33       31                                        Agglutination                                                                           None    None     None     None                                      Round Cells                                                                             1.8     8.3      0.2      0.2                                       ______________________________________                                    

                  TABLE 3                                                         ______________________________________                                        Original versus Separation Results for Patient JM                                                        Cell-Strainer                                                                          Cell-Strainer                                       Original                                                                              Swim-up  Method   Method                                    Patient JM                                                                              Semen   Method   (Vertical)                                                                             (Horizontal)                              ______________________________________                                        Volume (ml)                                                                             2.5     0.6      0.6      0.6                                       Sperm count/ml                                                                          56      47       51       36                                        (million)                                                                     Motility %                                                                              61      71       77       75                                        Progression                                                                             3       4        4        4                                         Velocity  28      32       31       33                                        Agglutination                                                                           None    None     None     None                                      Round Cells                                                                             0.7     0.3      0.2      0.1                                       ______________________________________                                    

                  TABLE 4                                                         ______________________________________                                        Original versus Separation Results for Patient FJ                                                        Cell-Strainer                                                                          Cell-Strainer                                       Original                                                                              Swim-up  Method   Method                                    Patient FJ                                                                              Semen   Method   (Vertical)                                                                             (Horizontal)                              ______________________________________                                        Volume (ml)                                                                             2.0     0.5      0.5      0.5                                       Sperm count/ml                                                                          18      14       14       12                                        (million)                                                                     Motility %                                                                              68      85       87       89                                        Progression                                                                             3       4        4        4                                         Velocity  21      30       31       31                                        Agglutination                                                                           None    None     None     None                                      Round Cells                                                                             0.9     0.1      0.1      0.1                                       ______________________________________                                    

It will be readily apparent that all three methods result in reductionsin total sperm count, however the total is still very high, viewedstatistically, relative to the likelihood of achieving fertilization.More importantly, in ail cases the motility percentage is increasedsignificantly. As is illustrated, in five out of six cases, theinvention provided greater improvement in motility than the Swim-upmethod of the prior art.

The present invention is for a simple, inexpensive and disposable systemfor extracting motile sperm and preventing inferior sperm from enteringthe reproduction process. Artificial reproduction methods are costly andtime-consuming procedures, and carry highly emotional overtones uponfailure. It is important to increase the likelihood of successfulreproduction, and the filtering of sperm is common in the industry. Theprior methods of filtering out the motile sperm involved centrifuges andswim-up methods to extract the motile sperm. These methods aretime-consuming and may lead to a low recovery of motile sperm, requiringadditional equipment and steps in the fertilization process. Theadditional equipment and time lead to a higher overall cost.

Yet an additional object of the invention is to provide a membraneapparatus that completely encloses the sample sperm before placing theenclosed sample into the container medium. This `tea bag` approachprovides a maximum surface area for the motile sperm to disperse fromthe sample into the medium.

A further object is to provide a sperm strainer system that can be usedfor animals as well as humans. In the preferred embodiment, human spermis considered, however the invention would work equally well with othersperm without deviating from the scope of the invention. With othertypes of sperm different sizes of mesh, and different mediums would beappropriate.

It is an object of the invention to provide a complete and portablesperm separation system. The invention can be packaged as a kit, andallow a fast and easy separation of the motile sperm. The kit would benot only cost effective, but can be used on locations that lack accessto additional laboratory equipment required by the prior methods.

An additional object of the invention is to provide a sperm strainerapparatus for separating motile sperm from a sperm sample, where thesperm strainer apparatus comprises a container with a disposed medium, amembrane assembly, wherein the membrane assembly has a nucleoporemembrane for passively separating the motile sperm from the spermsample, and wherein the membrane assembly engages the disposed medium.

Yet a further object of the invention is to provide a sperm strainerapparatus having a lid attachable to the container.

An additional object of the invention is to provide a sperm strainerapparatus where the container has a threaded portion for engaging amating threaded portion of the membrane assembly.

An object of the invention further includes a sperm strainer apparatuswhere the nucleopore membrane is 5-8 microns.

And a further object of the invention provides a sperm strainerapparatus wherein the nucleopore membrane is vertically disposed withinthe container.

Further object where a sperm fractionation apparatus having a membraneassembly that fits within the container and supports the nucleoporemembrane at an angle relative to the bottom surface of the container andextending downward substantially to the bottom surface of the container.

An object where a sperm fractionation apparatus has a membrane assemblythat fits within the container and rests on a plurality of stand-offsthat create a gap between a bottom surface of the container and a bottomsurface of the membrane assembly.

Further object, a method of separating motile sperm from a sperm samplecomprising the steps of contacting the sperm sample to a first surfaceof a nucleopore membrane having pores of 5-8 microns in diameter throughwhich motile sperm can generally migrate freely, contacting a receivingmedium suitable for sustaining live sperm to the opposing surface of thenucleopore membrane, incubating the sperm sample and receiving medium incontact with the membrane at ambient pressure and absent application ofcentrifugal effects until a sufficient population of sperm have migratedto the receiving medium.

Additional object where a method o f separating motile sperm utilizingan apparatus comprising a container and a membrane assembly, thenucleopore membrane being supported by the membrane assembly, thecontainer being configured to be divided into two chambers by themembrane assembly wherein the receiving medium can be contained in afirst chamber in contact with the first surface of the nucleoporemembrane and the sperm sample can be contained in a second chamber incontact with the second surface of the nucleopore membrane.

Further object a method of separating motile sperm, wherein theincubating is conducted for approximately 30 minutes.

Object where, a method of separating motile sperm, wherein theincubating is conducted at a temperature of approximately 37° C.

Object where a kit for the fractionation of motile sperm from a spermsample by incubation at ambient pressure without application ofcentrifugal effects comprising, a container; a receiving medium; amembrane assembly configured with a nucleopore membrane having pores of5-8 microns diameter, the container being divisible into two chambers byinsertion of the membrane assembly wherein the receiving medium can becontained in a first chamber in contact with the first surface of thenucleopore membrane and the sperm sample can be contained in a secondchamber in contact with the second surface of the nucleopore membrane,means for inserting and removing the receiving medium, and means forinserting and removing the sperm sample.

Object where a kit for the fractionation of motile sperm, furthercomprising a lid for covering the container.

Additional object where a kit for the fractionation of motile sperm,said means for inserting and removing saithed receiving mediumcomprising a syringe for displacing the medium.

And yet a further object includes kit for the fractionation of motilesperm the means for inserting and removing said sperm sample comprisinga pipette

Still other objects and advantages of the present invention will becomereadily apparent to those skilled in this art from the followingdetailed description, wherein I have shown and described only apreferred embodiment of the invention, simply by way of illustration ofthe best mode contemplated by me on carrying out my invention.

Other objects, features and advantages are apparent from thedescriptions presented in conjunction with the accompanying drawings.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 shows the disposable syringe with a needle dispensing medium intoa container;

FIG. 2 shows the container and membrane assembly, and a plastic pipetteplacing the semen sample into the membrane assembly;

FIG. 3 is an illustration of the incubation process;

FIG. 4 illustrates the ease and convenience of the clean-up process bydiscarding the cell strainer unit;

FIG. 5 depicts a syringe removing the motile sperm from the container;

FIG. 6(a) shows a container;

FIG. 6(b) shows a membrane assembly;

FIG. 6(c) shows the membrane assembly placed in the container;

FIG. 7(a) shows a container with a track member,

FIG. 7(b) shows an exploded view of the vertical membrane assembly;

FIG. 7(c) shows the vertical membrane assembly placed in the container;

FIG. 8(a) shows a container;

FIG. 8(b) shows a membrane assembly with standoffs;

FIG. 8(c) shows the membrane assembly placed in the container;

FIG. 9(a) shows a container with a lid;

FIG 9(b) shows a side view of the container and membrane assembly with alid.

DESCRIPTION OF THE PREFERRED EMBODIMENT

To those skilled in the art, the invention admits of many variations. Anenabling description of the preferred embodiment follows.

Referring to FIG. 1, a 1 ml disposable syringe 10 is used to placeapproximately 1-1.5 ml of a desired medium 20 into a container or dish30. There are various types of mediums commonly used in the industry,and the present invention would work with any of the mediums. The exactamount of medium is not important, although the rate of diffusion may belower for lesser amounts. It is also possible to use the presentinvention with minimal target medium to obtain an undiluted spermspecimen.

The size and shape of the container can vary, although the smaller thecontainer the more efficient and less costly the process. Plastic is thepreferred construction material in order to reduce costs, but glass isalso possible. In the preferred embodiment, the upper portion of thecontainer or dish has a threaded outer surface for engaging andretaining the membrane assembly. However, it is within the scope of theinvention to use other engaging and retaining methods such as innerthreads, snaps, and press-fit closing devices.

In FIG. 2, a disposable plastic pipette 40 is used to place the semensample 70 into the sperm membrane assembly 50. in the preferredembodiment, 1-1.5 ml of sperm laden source or sample material 70 isplaced into the unit. In practicing the invention, larger or smallercontainers and corresponding membrane units or strainer devices may beutilized and still be within the scope of the invention. These othersizes allow for different amounts of sample and target medium,separating membrane surface area, and sperm, in order to maximize thetransfer of the motile sperm for the particular purpose and the mediumused.

In the preferred embodiment, the unit has a lid attachment 60 thatcloses onto the mesh membrane. It prevents the mesh from falling intothe dish 30, aids in maintaining the correct temperature, and preventsparticles from falling into the strainer unit. The lid is not arequirement relative to the functioning of the mesh for separation, andother methods can be used to retain the mesh on the strainer withoutusing the lid.

The nylon mesh in the preferred embodiment has a pore size of 5-8microns, although variations are within the scope of the invention. Afiner pore size would further restrict the sperm and yield even a higherquality sperm but reduce the sample size. A greater pore size wouldallow more sperm to pass into the dish, but introduce less motile sperm.Non-human sperm samples may require different pore sizes. Nylon mesh isused for membrane of the present invention, although the invention worksequally well with other mesh materials.

FIG. 3 depicts the incubation process, wherein the motile sperm 80migrate through the mesh barrier and enter into the target section. Asshown, the cell strainer unit with the lid 60 closed is screwed onto thedish, and the sperm swim or migrate about in the sample or sourcesection until they encounter and penetrate the mesh barrier membraneinto the target medium.

In the preferred embodiment, the incubation process lasts thirty minutesat 37° C., although it depends on many variables. The amount of themedium, the surface area and pore size of the mesh, the temperature, theamount and density of sperm in the medium, and the mobility of the spermare all factors that should be considered in determining the incubationperiod.

At the end of the incubation period, the membrane apparatus 50 isremoved from the dish with the remaining portion of the original sample,as shown in FIG. 4. The membrane apparatus 50 with the remainingoriginal sample can be closed with the lid and disposed ofappropriately. The membrane apparatus can be easily discarded as shownin FIG. 4, eliminating messy clean-up.

As shown in FIG. 5, the target medium 90 now contains the recoveredsperm, ready for fertilization purposes. The dish with the threadedportion containing the refined sperm specimen can be closed with a lidand conveniently transported, or the specimen can easily be extractedusing a syringe and immediately used.

FIGS. 6(a), (b), and (c) illustrate an alternate embodiment of theinvention. An outer cell 100 is preferably made of plastic and conicalin design. An inner cell 110 is also made of plastic, and conical indesign. The inner cell 110 has a slightly smaller radius than the outercell 100, so that the inner cell 110 can fit within the outer cell 100.The bottom surface of the inner cell 110 has a nucleopore membrane 120with a pore size of between 5-8 microns. This is the optimal pore sizethat allows human motile sperm to penetrate, while leaving the dead orless motile sperm behind. Other pore sizes are within the scope of theinvention to accommodate other sperm sizes as for animals other thanhumans.

In this embodiment, the bottom surface membrane of the inner cell iselliptical in shape to provide greater surface area, and arranged at anangle that divides the volume in the lower region of the outer celldiagonally into a source volume and a target volume.

In operation, a suitable receiving medium is placed in the target volumearea, as shown in FIGS. 6(a) and (b). The inner cell 110 is placedinside the outer cell 100 such the lower end of the slanted bottomsurface membrane extends substantially down to the inner bottom surfaceof the outer cell, and that ail or substantially all of the bottomsurface membrane is wetted by the receiving medium. The sperm sample isplaced in the inner cell on the nucleopore membrane and should coversubstantially all of the membrane to assure the largest transfer surfacearea between the two mediums. A lid 130 is placed onto the cells and thesystem is allowed to incubate for about thirty minutes at approximately37° C.

Another embodiment uses a conical container that has a grooved innersurface 160 that divides the interior volume vertically into two halves,as shown in FIGS. 7(a), (b), and (c). A framed membrane section 170 thathas a 5-8 micron nucleopore membrane is inserted into groove 160 of thecontainer. This mating groove helps retain a tight fit against the innerwall of the container. The groove is not a necessary component, as theframed section can be placed into a container having no groove, as longas the fit assures there will be no leakage. The framed member section170 can have rubber, elastic, or other soft outer perimeter to retain atight fit on the inside wall of the container. A lid 200 with a lowerside groove can also aid in keeping the framed membrane in place.

In operation as shown in FIG. 7(c), the vertical separation scheme isillustrated. The framed membrane creates two compartments in thecontainer, a sample side and a receiving side. A buffer medium is addedto the receiving side, and a sperm sample is introduced into the sampleside. The stronger motile sperm migrate through the nucleopore membranein the manner of the invention, populating the receiving side with arelatively high motility sperm set.

FIGS. 8(a), (b), and (c) illustrate a further variation, where the innercontainer has standoffs 175 on the bottom surface. The standoffs keepthe inner container a small distance from the bottom surface of theoutside container to allow the sperm to settle and accumulate in thisregion. The standoffs can be on the inner container or the outercontainer, as long as it keeps the bottom surface of the inner containera small distance from the bottom surface of the outer container.

The lid 130 is made to cover the inner and outer cells and preventforeign debris from entering the cells, as shown in FIGS. 9(a) and (b).The lid 130 can be designed with a small slit 140 to provide ventilationand allow the cover to be removed with simple instruments. A handle canalso be installed to allow easy access. The outer perimeter of the lidcan envelop the outer circumference of the outer container 100, or beretained within the outer or inner containers and rest on the topsurface of the container. A notched perimeter allows for better seatingof the lid and is within the scope of the invention.

As will be realized, the invention is capable of other and differentembodiments and its several details are capable of modifications invarious obvious respects, all without departing from the invention.

In summary, the invention is distinguished from the prior art byproviding a passive separation of the motile sperm in a disposable andconvenient system. The system is cost effective as compared to existingmethods and allows on-site preparation of motile sperm forfertilization. It is less time consuming than other processes, and doesnot require extra equipment or taking the sample to a separate lab.

The foregoing description of the preferred embodiment of the inventionhas been presented for the purpose of illustration and description. Itis not intended to be exhaustive or to limit the invention to theprecise form disclosed. Many modifications and variations are possiblein light of the above teachings. It is intended that the scope of theinvention be limited not by this detailed description, but rather by theclaims appended hereto.

The invention is susceptible of many variations, all within the scope ofthe claims. The preferred embodiment described here and illustrated inthe figures should not be construed as in any way limiting.

No warranty is expressed or implied as to the actual degree of safety,security or support of any particular specimen of the invention in wholeor in part, due to differences in actual production designs, materialsand use of the products of the invention.

We claim:
 1. A sperm fractionation apparatus for separating motile spermfrom a sperm sample comprising:a nucleopore membrane with first andsecond opposing surfaces through which motile sperm can generallymigrate freely; a receiving medium suitable for sustaining live sperm;and a means for contacting said receiving medium with said first surfaceof said membrane and said sperm sample with said second surface of saidmembrane, thereby creating a transfer path through which migratingmotile sperm can pass from said sample to said receiving medium.
 2. Thesperm fractionation apparatus of claim 1, said means for contactingcomprising a container and a membrane assembly, said nucleopore membranebeing supported by said membrane assembly, said container beingconfigured to be divided into two chambers by said membrane assemblywherein said receiving medium can be contained in a first chamber incontact with said first surface of said nucleopore membrane and saidsperm sample can be contained in a second chamber in contact with saidsecond surface of said nucleopore membrane.
 3. The sperm fractionationapparatus according to claim 2, further comprising a lid attachable tosaid container.
 4. The sperm fractionation apparatus according to claim2, wherein the container has a threaded portion for engaging a matingthreaded portion of said membrane assembly.
 5. A sperm fractionationapparatus according to claim 1, wherein the pores of said nucleoporemembrane are 5-8 microns in diameter.
 6. A sperm fractionation apparatusaccording to claim 2, wherein said nucleopore membrane is verticallydisposed within said container.
 7. A sperm fractionation apparatusaccording to claim 2, wherein said membrane assembly fits within saidcontainer and supports said nucleopore membrane at an angle relative tosaid bottom surface of said container and extending downwardsubstantially to said bottom surface of said container.
 8. A spermfractionation apparatus according to claim 2, wherein said membraneassembly fits within said container and rests on a plurality ofstand-offs that create a gap between a bottom surface of said containerand a bottom surface of said membrane assembly.
 9. A method ofseparating motile sperm from a sperm sample comprising thesteps:contacting said sperm sample to a first surface of a nucleoporemembrane having pores of 5-8 microns in diameter through which motilesperm can generally migrate freely, contacting a receiving mediumsuitable for sustaining live sperm to the opposing surface of saidnucleopore membrane, incubating said sperm sample and receiving mediumin contact with said membrane at ambient pressure and absent applicationof centrifugal effects until a sufficient population of sperm havemigrated to said receiving medium.
 10. A method of separating motilesperm according to claim 9, said method utilizing an apparatuscomprising a container and a membrane assembly, said nucleopore membranebeing supported by said membrane assembly, said container beingconfigured to be divided into two chambers by said membrane assemblywherein said receiving medium can be contained in a first chamber incontact with said first surface of said nucleopore membrane and saidsperm sample can be contained in a second chamber in contact with saidsecond surface of said nucleopore membrane.
 11. A method of separatingmotile sperm according to claim 10, wherein said incubating is conductedfor approximately 30 minutes.
 12. A method of separating motile spermaccording to claim 10, wherein said incubating is conducted at atemperature of approximately 37° C.
 13. A kit for the fractionation ofmotile sperm from a sperm sample by incubation at ambient pressurewithout application of centrifugal effects comprising:a container; areceiving medium; a membrane assembly configured with a nucleoporemembrane having pores of 5-8 microns diameter, said container beingdivisible into two chambers by insertion of said membrane assemblywherein said receiving medium can be contained in a first chamber incontact with said first surface of said nucleopore membrane and saidsperm sample can be contained in a second chamber in contact with saidsecond surface of said nucleopore membrane, means for inserting andremoving said receiving medium, and means for inserting and removingsaid sperm sample.
 14. A kit for the fractionation of motile spermaccording to claim 13, further comprising a lid for covering saidcontainer.
 15. A kit for the fractionation of motile sperm according toclaim 13, said means for inserting and removing said receiving mediumcomprising a syringe for displacing said medium.
 16. A kit for thefractionation of motile sperm according to claim 13, said means forinserting and removing said sperm sample comprising a pipette.